| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
Department of Microbiology, University of Pennsylvania Medical School, Philadelphia, PA 19104
 |
Abstract |
|---|
 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
at levels similar to CD4+ T cells induced to
endogenous listerial Ags. These studies suggest that chromosomally
modified strains of L. monocytogenes may be useful as
vaccine vectors for the induction of Th1 T cell responses against
HIV. |
Introduction |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Listeria monocytogenes is a Gram-positive facultative
intracellular bacterium that enters the macrophage upon phagocytosis.
Its unusual ability to escape from the phagolysosome and live in the
cytoplasm of the cell (8), thus inducing both
CD8+ CTL and CD4+ T cells
(9, 10), has resulted in close investigation of
this organism as a vaccine vector to induce cell-mediated
immunity (11, 12). In addition to the production of a
strong CTL response to Ags delivered by L. monocytogenes
(13, 14, 15, 16), there is evidence that the
CD4+ T cell response to listerial Ags is of the
Th1 phenotype. Macrophages infected with L. monocytogenes
have been shown to produce IL-12 in vitro and to direct Th1 development
through the induction of IFN- (17, 18). In addition,
IL-12 has been shown to be produced in vivo after immunization with
live L. monocytogenes (19), and neutralization
of this cytokine results in exacerbated infection (20, 21). This effect can be reversed by treatment with IFN-
(21). Moreover, IFN- has been shown to be a critical
parameter of protection against L. monocytogenes in mice
with disrupted IFN- or IFN- receptor genes (22, 23).
On the other hand, neutralization of Th2 cytokines such as IL-10 and
IL-4 has been shown to increase resistance to L.
monocytogenes (24, 25). The concept that L.
monocytogenes-induced CD4+ T cells are
restricted to the Th1 phenotype may prove to be of importance in
vaccine development if the balance of Th1/Th2
CD4+ subsets is shown to be relevant to disease
progression in AIDS. In this study, therefore, we have investigated the
strength and phenotype of the CD4+ T cell
response to a foreign HIV gene product delivered by this potential
vaccine vector.
We have previously described a genetic method to permanently modify the
chromosome of L. monocytogenes so that HIV gene products can
be expressed as secreted proteins under the control of a copy of the
strong promoter of the hemolysin (hly) gene, which encodes
listeriolysin-O (26). Furthermore, we demonstrated that
mice immunized with one of these constructs, Lm-Gag, mount a strong,
specific, long-lasting CTL response against the HIV-1 Gag protein, and
that the response is directed predominantly against an epitope present
in the p24 portion of the protein (26, 27). In the present
work, we show the production in both BALB/c and C57BL/6 mice of
CD4+ T cells of a Th1 phenotype that peaks at day
7 after infection. Levels of IFN- produced by Gag-specific cells are
similar between the two strains. We also compared the production of
IFN- by Gag-specific CD4+ T cells induced
by Lm-Gag3 with that produced by CD4+ T cells
specific for all endogenous soluble secreted listerial Ags (SLA),
induced by both Lm-Gag and wild-type L. monocytogenes.
Levels of IFN- against the foreign HIV gene product were equivalent
to levels produced in response to the endogenous Ags delivered by
Lm-Gag and wild-type L. monocytogenes.
Finally,3 we have identified epitopes present
in the p24 portion of the protein in both strains of mice and found
that two of these epitopes are common to the H-2b
and H-2d MHC class II molecules. We discuss the
relevance of these findings with respect to the development of L.
monocytogenes as a vaccine against HIV-1.
|
Materials and Methods |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Female BALB/c (H-2d) and C57BL/6 (H-2b) were purchased from Charles River Laboratories (Raleigh, NC) and maintained in a pathogen-free microisolator environment. Mice used in this study were 6–12 wk old.
Bacterial strains
L. monocytogenes (Lm-wt) strain 10403S
(10) was the wild-type organism used in these studies. It
has an LD50 of 
5 x
104 when injected i.v. or i.p. into BALB/c mice.
Lm-Gag refers to either one of two recombinant strains of L.
monocytogenes, each of which carries a copy of the HIV-1 strain
HXB gag gene stably integrated into the listerial chromosome
and both of which secrete the gag gene product as determined
by Western blotting of secreted proteins (26) (M. Mata, A.
Zubair, Z.-J. Yao, K. Syres, and Y. Paterson, manuscript in
preparation). The two strains have an LD50 of
5 x 106 and 5 x
107. All strains were grown in brain/heart
infusion (BHI) medium (Difco, Detroit, MI).
Antigens
SLA is a mixture of secreted products of L. monocytogenes grown in vitro in BHI and may include such virulence factors as hemolysin, p60, phospholipase C, metalloprotease, and others. It was prepared by growing 10403S in BHI until it reached stationary phase. The bacteria were spun out and the supernatant was filtered. Ammonium sulfate was added to 80% saturation, and precipitated protein was sedimented by high-speed centrifugation. The pellet was resuspended in PBS and dialyzed against PBS to eliminate remaining ammonium sulfate.
Gag p24 was purified from an Escherichia coli heat-induced expression system, as previously described (28). In brief, E. coli was grown to log phase at 32°C in LB broth (Difco), and the temperature was raised to 42°C to induce production of p24. After 90 min of shaking, E. coli was pelleted and lysed, and inclusion bodies were purified. Inclusion bodies were then washed to eliminate any detergents remaining and suspended in 8 M urea. This preparation was further purified by anion-exchange purification (Gradifrac, Pharmacia Biotech, Piscataway, NJ) using SP and Q columns. Purity of p24 was then verified by Coomassie blue staining of SDS-PAGE gels as well as by Western blot (data not shown).
SIV-Nef was purified from an E. coli isopropyl
-D-thiogalactoside induction system (kind gift
of Dr. Casey Morrow, University of Alabama, Birmingham, AL). In brief,
E. coli was grown to log phase in LB broth (Difco) with
carbenicillin and kanamycin (Sigma, St. Louis, MO) at 37°C. Isopropyl
-D-thiogalactoside was added at a final
concentration of 0.5 µM, and cells were incubated an additional
2 h at 30°C. After overnight storage at room temperature,
E. coli was pelleted and sonicated. The resulting protein
solution was mixed with 50% slurry of glutathione-Sepharose 4B beads
(Pharmacia Biotech) and rocked for 30 min at 4°C. After washing the
matrix with thrombin cleavage buffer (Novagen, Madison, WI), the
protein was cleaved using thrombin protease (Novagen), collected, and
dialyzed against PBS buffer. Purity of Nef was verified by Coomassie
blue staining of SDS-PAGE gels as well as by Western blot (data not
shown).
Twenty-two overlapping 20-mer peptides (Table I
) with 10-aa overlaps, spanning residues
133–362 of the HIV-1SF2 Gag protein encompassing the p24 region, were
provided by the MRC AIDS reagent project, London, U.K. (Repository
reference: ADP 788.1–22).
|
Proliferation assays
Six- to twelve-week-old female mice were immunized i.p. with 106 or 107 live Lm-Gag, or with 104 live 10403S (Lm-wt). Splenocytes were obtained from mice 1 wk after immunization, and T cells were purified by nylon wool purification, as previously described (29). Briefly, splenocytes were added to a nylon wool column to eliminate adherent cells and were incubated at 37°C for 45 min, eluted, and washed. This step was performed twice for higher purity. A total of 5 x 105 nylon wool purified splenocytes was cultured in triplicate in 96-well microplates with Ag in 200 µl of RPMI 1640 supplemented with 10% FBS, 100 U/ml penicillin, 100 µg/ml streptomycin, 2 mM L-glutamine, and 50 µM 2-ME (RP-10). An equal number of irradiated (3000 rad) splenocytes from naive mice were added as APCs. T cells and APCs were cocultured for 3 days at 37°C. After 48 h, [3H]thymidine at 50 µCi/ml was added to the cultures for 16–24 h. Plates were harvested on day 3 using a Tomtec harvester 96 (Hamden, CT), and [3H]thymidine incorporation was measured with a MicroBeta Trilux scintillation counter (Wallac, EG&G, Gaithersburg, MD).
Complement-mediated depletion of CD4+ and CD8+ T cells
Nylon wool-purified splenocytes were resuspended in RPMI 1640 media supplemented with 5% FCS at a concentration of 1 x 107 cells/ml. Supernatants from hybridoma 2.43 (anti-CD8) or hybridoma GK1.5 (anti-CD4) containing the respective Abs were added to the cells at a 1/2 dilution. Cells and Abs were incubated for 30 min at 4°C. After this time, cells were spun and resuspended at the same concentration, and rabbit complement H-2 (PelFreez Clinical Systems, Brown Deer, WI) was added at a 1/5 dilution. Cells and complement were incubated for 45 min at 37°C. After this time, cells were washed, counted, and resuspended in RP-10 and used in a proliferation assay, as described above.
ELISA cultures
Six- to eight-week-old female mice were immunized by i.p. inoculation with 106 or 107 live Lm-Gag, or with 104 live Lm-wt. Splenocytes (5 x 105 cells/ml), obtained from mice at several different time points after immunization, were cultured in 24-well plates with Ag at 37°C in 1 ml of RPMI 1640 supplemented with 10% FBS, 100 U/ml penicillin, 100 µg/ml streptomycin, 2 mM L-glutamine, and 50 µM 2-ME. After 3 days, supernatants of duplicate cultures were collected and stored at -20°C until samples were tested for cytokines by ELISA.
Cytokine ELISAs
IFN- and IL-4 were measured using a specific sandwich ELISA,
as described previously (30, 31, 32). IFN- was detected
using the mAb R46-A2 at 5 µg/ml and polyclonal rabbit anti-IFN-
used at an optimal dilution (kindly provided by Dr. Phillip Scott,
University of Pennsylvania, Philadelphia, PA). The levels of IFN-
were calculated by comparison with a standard curve using murine
rIFN- (Life Technologies, Gaithersburg, MD). IL-4 was detected using
mAb 11B11 at 1 µg/ml and biotinylated mAb BVD-6 used at an optimal
dilution. The levels of IL-4 were calculated by comparison with a
standard curve using supernatants from the IL-4-secreting X-4 cell line
(33). Plates were developed using a peroxidase-conjugated
goat anti-rabbit IgG Ab (IFN-) or peroxidase-conjugated
streptavidin (IL-4) (Jackson ImmunoResearch, West Grove, PA) and ABTS
(2.2'-azino-di[3-ethyl-benzthiazoline sulfonate(6)] as a
peroxidase substrate (Kirkegaard & Perry Laboratories, Gaithersburg,
MD). Plates were then read at 405 nm. The lower limit of detection for
the assays was as follows: 30 pg/ml for IFN- and 0.2 U/ml
for IL-4.
|
Results |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
To study the CD4+ T cell response to
HIV-Gag, we first measured the ability of Lm-Gag immune splenocytes to
proliferate in response to exogenous HIV-Gag Ag. We used splenocytes
from BALB/c mice immunized with Lm-wt or the Lm-Gag recombinant strains
and stimulated them with Ag for 3 days before pulsing with
[3H]thymidine. Whereas splenocytes from both
groups of mice were able to proliferate equally well in response to SLA
(total secreted proteins from L. monocytogenes), only
splenocytes from mice immunized with Lm-Gag proliferated in response to
the in vitro addition of purified p24, a proteolytic product of the Gag
protein (Fig. 1). The response to Gag p24
was specific because splenocytes from Lm-Gag-immunized mice were not
stimulated by rSIV239-Nef produced and purified
by a similar protocol (data not shown). Not unexpectedly, proliferation
of Lm-Gag splenocytes to p24 was not as great as to SLA, which includes
numerous Ags secreted by L. monocytogenes, including a
number of known virulence factors (34).
|
To further characterize the proliferative response to p24 Gag, we
wished to identify epitopes in two different strains of mice, BALB/c
(H-2d) and C57BL/6 (H-2b).
Twenty-two overlapping peptides (Table I
) that span the entire p24
region of the Gag protein were used as Ags in proliferation assays with
splenocytes from mice immunized with Lm-Gag. We were able to identify
two and three epitopes for the H-2d and
H-2b haplotypes, respectively (Fig. 2, A and B). In
addition, none of the peptides induced proliferative responses in
splenocytes from mice immunized with the wild-type strain of L.
monocytogenes (data not shown). Interestingly, two of these
epitopes were common for both strains of mice. Promiscuous epitopes
that bind to more than one haplotype have previously been identified
for HIV-Env (35, 36). Indeed, such epitopes are considered
advantageous for peptide-based vaccine approaches because they can be
used to induce immunity in outbred populations (37, 38).
To determine whether the common epitopes were immunodominant for both
haplotypes, proliferation assays were performed at limiting dilutions
(Fig. 3, A and B).
The data show that although two peptides are recognized by both strains
of mice, namely MRC-2 (Gag143–162) and MRC-17
(Gag193–212), only MRC-2 is immunodominant and
solely for the H-2b haplotype (Fig. 3B). In BALB/c mice, MRC-13
(Gag253–272) is immunodominant for
H-2d (Fig. 3A).
|
|
,
depletion of the CD4+ population, but not the
CD8+ population, resulted in total abrogation of
the proliferative response, confirming that this is, indeed, a measure
of the CD4+ T cell response. Depletion of
CD8+ T cells did result in diminished stimulation
index values, suggesting a possible contribution by this T cell subset.
This difference may be due to an increased background proliferative
response of the CD4+ T cell population remaining
after CD8+ T cell depletion, which was observed.
It is also possible that activated CD8+ T cells
may be playing a role by producing cytokines that could enhance the
CD4+ T cell proliferative response.
|
A major incentive for using L. monocytogenes as a
vaccine vector for HIV is its ability to induce a strong Th1 phenotype
response. Although it seems likely that the CD4+
T cell response to a foreign protein secreted by L.
monocytogenes would have a similar phenotype to that specific for
endogenous listerial Ags, this has not previously been investigated. To
verify that the Gag-specific response is also of the Th1 phenotype, we
examined the cytokine profile of Gag-specific T cells in vitro.
Splenocytes from mice immunized with Lm-Gag were cultured with Ag for 3
days. Supernatants from these cultures were then tested for IFN- and
IL-4 using specific two-site or sandwich ELISAs. As shown in Fig. 5, while splenocytes from both Lm-wt- and
Lm-Gag-immunized mice produced IFN- in response to SLA, only
splenocytes from Lm-Gag-immunized mice responded to p24 by producing
IFN-. IL-4 production was below the level of detection of this assay
(0.2 U/ml) (data not shown).
|
peaked at day 7 for both p24 and SLA
responses consistent with adaptive immune cells as the source of
cytokine secretion. Both CD4+ and
CD8+ T cells are known to produce IFN- in an
Ag-specific manner in listeriosis as well as in viral diseases
(39, 40). The exogenous p24 used as a source of Ag in
these assays is unlikely to stimulate CD8+ T
cells. However, to determine the role played by
CD4+ and CD8+ T cells in
Ag-specific production of IFN-, we took advantage of the previous
identification of T cell epitopes. In the H-2b
haplotype, we could confirm that CD4+ T cells
that recognize the two epitopes identified (MRC-2 and MRC-17) produce
IFN- (Fig. 6B), and that
the level of production correlates with the antigenicity of the
epitope, as determined by proliferation assays (Fig. 3B).
The levels of IFN- production in this particular experiment were
lower than in other experiments presented in this work and are a good
example of the variability observed in these types of assays and the
difficulty of working with fresh splenocytes and in vivo systems
instead of cell lines and clones. Nonetheless, the patterns of
secretion are reproducible from one experiment to another. For T cells
of the H-2d haplotype, we were able to test the
role of both CD8+ and CD4+
T cells using the MHC class II epitopes identified above as well as a
MHC class I epitope previously described (27),
Gag197–205. Although some T cells clearly
respond to the MHC class I epitope with production of IFN-, the
level of production by T cells recognizing MRC-13, the MHC class II
epitope, is substantially higher (Fig. 6A). Again, a
comparison of the different CD4+ epitopes
corroborates immunogenicity of the epitopes determined by proliferation
assays (Fig. 3A).
|
|
Discussion |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Although differences in resistance to L. monocytogenes have
been reported among different strains of mice (49, 50),
these do not appear to be related to a predisposition by susceptible
strains to a Th2 CD4+ T cell phenotype, as is the
case in leishmaniasis (51). The immune response to
L. monocytogenes, in both susceptible and resistant strains,
is characterized by a strong Th1 cell-mediated response with the
production of high levels of IL-12, TNF-
, and IFN- and strong CTL
responses to several Listerial Ags (52). mRNA
levels of several cytokines, including IL-1, IL-6, GM-CSF, and
TNF-, can be detected in vivo, in a variety of strains of mice, as
early as 30 min after infection in the liver and spleen
(53, 54, 55, 56). IFN- mRNA can be detected as early as 30 min
in the liver, but a significant increase is seen after 16 h in the
spleen (55, 56). These data correlated with cytokine
secretion by splenocytes harvested 6 h after infection and
cultured for 48 h in the presence of Ag (56).
Furthermore, using in vivo Ab depletion or cytokine knockout mice, some
of these cytokines, namely IL-6 (57), TNF- (19, 58), IL-12 (20, 21), and IFN- (22, 23), have been shown to be indispensable for protection against
listerial infection. Although control of the infection appears to be
mediated by neutrophils (59) and activated macrophages
(60, 61, 62), the presence of T cells, in particular CTLs, is
necessary to obtain sterilizing immunity (52). In
addition, memory responses are clearly mediated by CD8 T cells in the
mouse model (63, 64).
Clearly, the outcome of HIV and listerial infections is very different in that the immune response to listerial Ags results in sterilizing immunity and protection from further challenge in mice, whereas the immune response to HIV infection is not always successful in controlling viral replication. There is, therefore, an urgent need to develop anti-HIV vaccines to induce immune responses that can control infection. To determine what the requirements are for these vaccines, studies have focused on the immune parameters of those individuals that have mounted effective immune responses against the virus, which were discussed above. Because the immune response to listerial Ags and the immune parameters thought to be required for effective HIV vaccine approaches have critical elements in common, we have been developing L. monocytogenes constructs expressing HIV Ags as possible AIDS vaccines (11, 26, 27) (M. Mata, A. Zubair, Z.-J. Yao, K. Syres, and Y. Paterson, manuscript in preparation). Using one of these constructs that expresses the gag gene, we were able to show the induction of strong CTL responses (26) and identified an epitope in the H-2d haplotype (27). In this study, we have examined the CD4+ T cell compartment in anti-HIV Gag immunity induced by Lm-Gag in two strains of mice that differ in both MHC haplotype and genetic background.
The first question we asked was whether we could detect proliferative
responses in vitro to recombinant p24, a proteolytic product of the Gag
protein that had been shown to contain the class I-restricted epitope
in BALB/c mice (26). A proliferative response to a large
protein in vitro is usually indicative of the presence of Ag-specific
CD4+ T cell responses. Our data, then, suggested
that the proliferative response seen in mice immunized with Lm-Gag, but
not with the wild-type strain of L. monocytogenes, was
CD4+ specific (Fig. 1). Furthermore, utilizing a
series of overlapping peptides that spanned the whole sequence of p24
(Table I), we were able to identify several epitopes in two different
haplotypes, namely H-2d and
H-2b (Fig. 2, A and B).
More interestingly, two of these epitopes were common to both strains
of mice.
A potential problem in vaccine approaches that focus on isolated Ags or
T cell peptide epitopes is the diversity of the MHC molecules in an
outbred population that could result in Ags being presented by some but
not other MHC haplotypes, leaving a segment of the population
unprotected (37, 38). Therefore, the identification of
epitopes that can bind to more than one haplotype was interesting.
However, the immunodominance patterns observed for the two strains of
mice indicated that although Gag293–312 (MRC-17)
and Gag143–162 (MRC-2) are recognized by both
strains, they are weak Ags for H-2d-restricted T
cells (Fig. 3).
A major objective in using L. monocytogenes as a vaccine
approach for HIV is to exploit its characteristic induction of a Th1
response. In this study, we have confirmed that the response to a
recombinant Ag delivered by L. monocytogenes showed this
particular cytokine pattern. Although splenocytes from mice immunized
with either the Lm-Gag or the wild-type strain responded to listerial
Ags (SLA), only mice immunized with Lm-Gag responded to the recombinant
p24 Ag in vitro by secreting IFN-, whereas IL-4 production was not
detectable in any of the groups. The IFN- response peaked at day 7,
correlating with the activation of T cells shown to peak at this time
point in a primary infection with L. monocytogenes
(65, 66). Other, non-T cell sources of IFN- such as NK
cells or 
T cells would be expected to feature more prominently
early in infection, nor are they known to be specific for conventional
protein Ags. In addition, using MHC-specific epitopes (Fig. 6), we were
able to verify that the major cellular source of this cytokine was,
indeed, a CD4+, MHC class II-restricted T
cell.
In this study, we have shown the induction of Gag-specific CD4+ T cells with a Th1 phenotype in response to HIV-Gag delivered by L. monocytogenes and demonstrated that such response is comparable with the response mounted to the endogenous listerial Ags. This response, in combination with the CD8+ T cell response reported in previous work (26, 27), encourages the further exploration of L. monocytogenes as a vaccine vector for HIV, including a comparison with other live vectors such as vaccinia virus, Salmonella sp., and bacillus Calmette-Guérin, as well as other approaches such as peptide vaccines, recombinant proteins, and DNA vaccines. In addition, the ability to use L. monocytogenes to deliver recombinant Ags with the consequent induction of both CD4+- and CD8+-specific responses may be useful for the prevention of other viral infections (11, 12, 14, 15, 16) and treatment of other diseases such as cancer (11).
|
Acknowledgments |
|---|
|
Footnotes |
|---|
2 Address correspondence and reprint requests to Dr. Yvonne Paterson, Department of Microbiology, 323 Johnson Pavilion, 3610 Hamilton Walk, University of Pennsylvania, Philadelphia, PA 19104-6076. E-mail address:
3 Abbreviations used in this paper: Lm-Gag, L. monocytogenes strain expressing HIVgag; BHI, brain/heart infusion; Lm-wt, L. monocytogenes wild-type strain; SLA, secreted listerial Ags.
Received for publication March 12, 1999. Accepted for publication May 13, 1999.
|
References |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Th2 switch is a critical step in the etiology of HIV infection. Immunol. Today 14:107.[Medline]
-TCR transgenic T cells is macrophage dependent. Int. Immunol. 5:371. production in response to live or dead bacteria by TNF and other factors. J. Immunol. 161:1447.. J. Immunol. 152:1883.[Abstract]
genes. Science 259:1739. receptor. Science 259:1742.. J. Immunol. Methods 104:137.[Medline]
interferon and tumor necrosis factor in host resistance against primary and secondary Listeria monocytogenes infections. Infect. Immun. 57:3331. and IL-4 responses to viral infections: requirements for IL-2. J. Immunol. 160:5007. receptor-deficient mice. J. Immunol. 158:5297.[Abstract]
This article has been cited by other articles:
|
|
 |
|
  M. G. Cottingham, A. van Maurik, M. Zago, A. T. Newton, R. J. Anderson, M. K. Howard, J. Schneider, and M. A. Skinner Different Levels of Immunogenicity of Two Strains of Fowlpox Virus as Recombinant Vaccine Vectors Eliciting T-Cell Responses in Heterologous Prime-Boost Vaccination Strategies. Clin. Vaccine Immunol., July 1, 2006; 13(7): 747 - 757. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 C. N. Kotton and E. L. Hohmann Enteric Pathogens as Vaccine Vectors for Foreign Antigen Delivery Infect. Immun., October 1, 2004; 72(10): 5535 - 5547. [Full Text] [PDF]
|
|
|
|
 |
|
 R. Stevens, K. E. Howard, S. Nordone, M. Burkhard, and G. A. Dean Oral Immunization with Recombinant Listeria monocytogenes Controls Virus Load after Vaginal Challenge with Feline Immunodeficiency Virus J. Virol., August 1, 2004; 78(15): 8210 - 8218. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. Giri, K. E. Ugen, and D. B. Weiner DNA Vaccines against Human Immunodeficiency Virus Type 1 in the Past Decade Clin. Microbiol. Rev., April 1, 2004; 17(2): 370 - 389. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 D. A. Sewell, D. Douven, Z.-K. Pan, A. Rodriguez, and Y. Paterson Regression of HPV-Positive Tumors Treated With a New Listeria monocytogenes Vaccine Arch Otolaryngol Head Neck Surg, January 1, 2004; 130(1): 92 - 97. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
H. Saklani-Jusforgues, E. Fontan, N. Soussi, G. Milon, and P. L. Goossens Enteral Immunization with Attenuated Recombinant Listeria monocytogenes as a Live Vaccine Vector: Organ-Dependent Dynamics of CD4 T Lymphocytes Reactive to a Leishmania major Tracer Epitope Infect. Immun., March 1, 2003; 71(3): 1083 - 1090. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
V. Novitsky, H. Cao, N. Rybak, P. Gilbert, M. F. McLane, S. Gaolekwe, T. Peter, I. Thior, T. Ndung'u, R. Marlink, et al. Magnitude and Frequency of Cytotoxic T-Lymphocyte Responses: Identification of Immunodominant Regions of Human Immunodeficiency Virus Type 1 Subtype C J. Virol., September 11, 2002; 76(20): 10155 - 10168. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
H. Angelakopoulos, K. Loock, D. M. Sisul, E. R. Jensen, J. F. Miller, and E. L. Hohmann Safety and Shedding of an Attenuated Strain of Listeria monocytogenes with a Deletion of actA/plcB in Adult Volunteers: a Dose Escalation Study of Oral Inoculation Infect. Immun., July 1, 2002; 70(7): 3592 - 3601. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. L. Marzo, V. Vezys, K. Williams, D. F. Tough, and L. Lefrancois Tissue-Level Regulation of Th1 and Th2 Primary and Memory CD4 T Cells in Response to Listeria Infection J. Immunol., May 1, 2002; 168(9): 4504 - 4510. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
G. R. Gunn, A. Zubair, C. Peters, Z.-K. Pan, T.-C. Wu, and Y. Paterson Two Listeria monocytogenes Vaccine Vectors That Express Different Molecular Forms of Human Papilloma Virus-16 (HPV-16) E7 Induce Qualitatively Different T Cell Immunity That Correlates with Their Ability to Induce Regression of Established Tumors Immortalized by HPV-16 J. Immunol., December 1, 2001; 167(11): 6471 - 6479. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. Hiroi, H. Goto, K. Someya, M. Yanagita, M. Honda, N. Yamanaka, and H. Kiyono HIV Mucosal Vaccine: Nasal Immunization with rBCG-V3J1 Induces a Long Term V3J1 Peptide-Specific Neutralizing Immunity in Th1- and Th2-Deficient Conditions J. Immunol., November 15, 2001; 167(10): 5862 - 5867. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. A. Vazquez-Boland, M. Kuhn, P. Berche, T. Chakraborty, G. Dominguez-Bernal, W. Goebel, B. Gonzalez-Zorn, J. Wehland, and J. Kreft Listeria Pathogenesis and Molecular Virulence Determinants Clin. Microbiol. Rev., July 1, 2001; 14(3): 584 - 640. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. V. Rayevskaya and F. R. Frankel Systemic Immunity and Mucosal Immunity Are Induced against Human Immunodeficiency Virus Gag Protein in Mice by a New Hyperattenuated Strain of Listeria monocytogenes J. Virol., March 15, 2001; 75(6): 2786 - 2791. [Abstract] [Full Text]
|
|
|
|
|
|
D. Weissman, H. Ni, D. Scales, A. Dude, J. Capodici, K. McGibney, A. Abdool, S. N. Isaacs, G. Cannon, and K. Kariko HIV Gag mRNA Transfection of Dendritic Cells (DC) Delivers Encoded Antigen to MHC Class I and II Molecules, Causes DC Maturation, and Induces a Potent Human In Vitro Primary Immune Response J. Immunol., October 15, 2000; 165(8): 4710 - 4717. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. E. Touloukian, W. W. Leitner, S. L. Topalian, Y. F. Li, P. F. Robbins, S. A. Rosenberg, and N. P. Restifo Identification of a MHC Class II-Restricted Human gp100 Epitope Using DR4-IE Transgenic Mice J. Immunol., April 1, 2000; 164(7): 3535 - 3542. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
N. Soussi, G. Milon, J.-H. Colle, E. Mougneau, N. Glaichenhaus, and P. L. Goossens Listeria monocytogenes as a Short-Lived Delivery System for the Induction of Type 1 Cell-Mediated Immunity against the p36/LACK Antigen of Leishmania major Infect. Immun., March 1, 2000; 68(3): 1498 - 1506. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
